Skip to main content
NCBI home page
Biology logoLink to Biology
. 2025 May 17;14(5):562. doi: 10.3390/biology14050562

The Coexistence of Bicellular and Tricellular Pollen Might Be the Third Type of Pollen Cell Number: Evidence from Annonaceae

Yangying Gan 1, Qi Zhang 2, Chunfen Xiao 3, Jingyao Ping 4,*
Editor: Bartosz Płachno
PMCID: PMC12109490  PMID: 40427751

Simple Summary

Anther is thought to release either bicellular or tricellular pollen when mature. In the present work, we found that 16 species from 10 genera of Annonaceae shed both bicellular and tricellular pollen. This is the first time that so many species with both types of pollen has been observed in the same family. Combined with reports from other families, the plants that were known to shed both types of pollen included 15 families, 40 genera, and 52 species. Our results indicate that the coexistence of bicellular and tricellular pollen might be the third type of pollen cell number. And the systematic relationship among them is needed to be reanalyzed.

Keywords: Annonaceae, pollen cell number, bicellular pollen, tricellular pollen

Abstract

Anther is thought to release either bicellular or tricellular pollen when mature. Though a few species had been found to shed both bicellular and tricellular pollen, due to their low frequency, they had been overlooked as special cases of bicellular or tricellular pollen in previous phylogenetic studies. In the present work, the pollen cytologies of 89 species from 26 genera of Annonaceae were observed using the overall transparency method and paraffin sectioning method. The results show that 73 species from 25 genera distribute bicellular pollen, while 16 species from 10 genera shed both bicellular and tricellular pollen. This is the first time that so many species with both types of pollen has been observed in the same family. Combined with reports from other families, the plants that were known to shed both types of pollen included 15 families, 40 genera, and 52 species. Our results indicate that the coexistence of bicellular and tricellular pollen might be the third type of pollen cell number. And the systematic relationship among them is needed to be reanalyzed.

1. Introduction

Most angiosperms contain only one vegetative cell and one reproductive cell in their pollen before dispersal, known as bicellular pollen. About 30% of angiosperms complete the second cell division before dispersal, forming two reproductive cells and one vegetative cell, known as tricellular pollen [1]. There has long been controversy regarding the systematic evolution of pollen cell numbers. Early researchers believed that bicellular pollen was primitive and that the evolution from bicellular to tricellular pollen was irreversible [2,3]. Based on these claims, Webster and Rupert proposed the “Schürhoff–Brewbaker Law” [4]. However, subsequent research has suggested that the evolution from bicellular to tricellular pollen is reversible, and is skeptical of the primitive trait [1]. Additionally, in all previous systematic analyses on pollen cell numbers, only bicellular and tricellular pollen were taken into consideration. Although a few species had been found to shed both types of pollen [5,6,7], due to their low frequency, they had been considered as special cases and excluded from samples [1,3]. But, over time, more and more species with both kinds of pollen have been discovered [8,9,10,11,12,13,14,15,16].

Annonaceae was reported to be bicellulate by Brewbaker [3], but the coexistence of bicellular and tricellular pollen has been found in anthers of Annona cherimola Mill. and Mitrephora macclurei [6,9]. According to previous reports, Annonaceae has great diversity in terms of pollen size, shape, polarity, symmetry, dispersal unit, number/position/shape of germination aperture, ornamentation, and tectal and infratectal characters [17,18,19,20]. As a large family, comprising 107 genera and c.2400 species [21], it may also be diverse in terms of pollen cell number. But, until now, only nine species from six genera are known to have bicellular pollen [1,3,22,23,24,25], and two species from two genera are known to have both types of pollen [6,9,26]. Among the other undetected species, will there be more abundant discoveries (more taxa with both types of pollen, or even tricellular pollen)? With these questions and expectations, we have observed the pollen cell numbers of most of the Annonaceae plants currently distributed or introduced in China. The results may enrich our botanical understanding of pollen cell numbers and provide more evidence for systematic evolution research.

2. Materials and Methods

2.1. Materials

Fully mature flowers from 89 species across 26 genera of Annonaceae were collected from 2019 to 2021, taking about 5–10 flowers per plant and 5–10 anthers per flower annually for 2–3 consecutive years. The sampling species information includes the location, introduction number, and specimen number. The introduction number is available on the official website of the three botanical gardens (Xishuangbanna Tropical Botanical Garden: https://www.xtbg.ac.cn/ (accessed on 15 March 2024); Wuhan Botanical Garden: http://www.whiob.ac.cn/ (accessed on 18 March 2024); South China Botanical Garden: https://www.scbg.ac.cn/ (accessed on 20 March 2024)). The specimen number is available on the official website of the National Plant Specimen Resource Center of China (http://www.nsii.org.cn (accessed on 22 March 2024)) or the Chinese Virtual Herbarium (https://www.cvh.ac.cn/ (accessed on 25 March 2024)). All materials were fixed in formalin acetic alcohol (FAA: 70% alcohol, formaldehyde, and glacial acetic acid in a ratio of 90:5:5).

2.2. Methods

The pollen cell numbers were observed either by the overall transparency method described by Fu et al. [27] or the paraffin sectioning method described by Gan and Xu [9]. The proportion of anthers with both types of pollen in the sample anthers was also calculated.

Overall transparency method: All pollen was peeled off from mature anthers under a dissecting microscope to create a pollen suspension. The suspension was then subjected to hydrochloric acid hydrolysis, hematoxylin staining, gradient alcohol dehydration (30%, 50%, 70%, 90%, 95%, 100%) and transparent treatment with methyl salicylate, followed by DAPI staining, and placed on a glass slide dripped with clove oil for sealing. Fluorescence microscopy was used for observation and photography.

Paraffin sectioning method: For 64 species that the overall transparency method was not applicable, the paraffin sectioning method described by Gan and Xu [9] was applied instead. FAA-fixed anthers were subjected to ethanol gradient dehydration, xylene transparency, safranin-fixed green staining, paraffin embedding, and sectioning for 9 μm. A Leica DFC550 optical microscope (Leica Microsystem, Wetzlar, Germany) was used for observation and photography.

3. Results

Through the observation of 89 species from 26 genera of Annonaceae, we found that 16 species from 10 genera disperse both types of pollen (Figure 1, Figure 2 and Figure 3, Table 1), while 73 species from 25 genera shed bicellular pollen (Figure 4, Figure 5, Figure 6 and Figure 7, Table 1). Among the 16 species that disperse both types of pollen, 12 of them had more than half of anthers that contain both types of pollen, and for others such as Uvaria grandiflora Roxb, Uvaria calamistrata Hance, Mitrephora wangii Hu and Artabotrys hexapetalus (L. f.) Bhandar, 20–30% of sample anthers generally had both types of pollen. Our findings increase the number of known plants in Annonaceae that disperse both types of pollen from 2 genera and 2 species to 10 genera and 17 species (Table 2), However, pure tricellular pollen has not yet been detected in this family. In this study, we also observed both types of pollen (Figure 2Q) in the anthers of Annona cherimola Mill., confirming the research report of Lora et al. [26]. Additionally, we once again observed both types of pollen within the same pollen unit (Figure 2I–L), supporting the report by Gan and Xu [9].

Figure 1.

Figure 1

Open in a new tab

Mixed developmental stages of Annonaceae pollen before anther release (I). (A–D) The bicellular (A,C,D left) and tricellular (B,D right) pollen of Uvaria grandiflora Roxb; (E–H) the bicellular (G) and tricellular (E,F,H) pollen of Uvaria kurzii (King) P. T. Li; (I–L) the bicellular (I–K) and tricellular (L) pollen of Artabotrys hexapetalus (L. f.) Bhandar; (M–P) the bicellular (N right, P) and tricellular (M,N left,O) pollen of Artabotrys hongkongensis Hance. Scale bar = 50 μm. Arrows show cell nucleus.

Figure 2.

Figure 2

Open in a new tab

Mixed developmental stages of Annonaceae pollen before anther release (II). (A–D) The bicellular (C left,D right) and tricellular (A,B,C right,D right) pollen of Artabotrys pachypetalus B. Xue & Junhao Chen; (E–H) the bicellular (E) and tricellular (F–H) pollen of Goniothalamus calvicarpus Craib; (I–L) the bicellular (I,J down,K,L) and tricellular (J up) pollen of Goniothalamus gardneri Hook. f. et Thoms; (M–P) the bicellular (N right, O) and tricellular (M,N left,P) pollen of Fissisfigma polyanthum Hook. f. et Thoms; (Q) the tricellular pollen of Annona cherimola Mill; (R,S) the bicellular (R) and tricellular (S) pollen of Uvaria calamistrata Hance. Scale bar = 50 μm. Arrows show cell nucleus.

Figure 3.

Figure 3

Open in a new tab

Mixed developmental stages of Annonaceae pollen before anther release (III). (A–D) The bicellular (A up,B down,C,D) and tricellular (A down,B up) pollen of Mitrephora wangii Hu; (E–H) the bicellular (F,G) and tricellular (E,H) pollen of Meiogyne oligocarpa B. Xue & Y. H. Ta; (I,J) the bicellular (J up) and tricellular (I,J down) pollen of Dasymaschalon rostratum Merr. & Chun; (K,L) the bicellular (K) and tricellular (L) pollen of Orophea laui Leonardía & Kessler; (M–P) the tricellular (M right,N) and bicellular (O,P) pollen of Polyalthia cheliensis Hu; (Q–T) the tricellular (Q,R) and bicellular (S,T) pollen of Goniothalamus chinensis Merr. et Chun. Scale bar = 50 μm. Arrows show cell nucleus.

Table 1.

List of investigated species with provenance, voucher number and corresponding figure plate.

No. Taxon Provenance a Voucher Figures
1 Desmos chinensis Lour. SCBG xx060308, 20081072, xx271155 Figure 4A
2 Desmos dumosus (roxb.)saff. XTBG 00012542, C06009, 273677 Figure 4B
3 Desmos yunnanensis (Hu) P. T. Li XTBG 275056, 258389 Figure 4C
4 Dasymaschalon trichophorum Merr. SCBG 20011172, 19975026, 19970018 Figure 4D
5 Dasymaschalon filipes (Ridl.) Ridl.Ban XTBG 1320030093, 0020220650 Figure 4E
6 Dasymaschalon rostratum Merr. & Chun * XTBG 0020023226, 0020020479 Figure 3I,J
7 Dasymaschalon macrocalyx Finet & Gagnep. XTBG 1320030078, 0020022105 Figure 4F
8 Polyalthia suberosa (Roxburgh) Thwaites SCBG 20010982, 20070804, 20140665 Figure 4G
9 Polyalthia cheliensis Hu * XTBG 0020081058, 0020100741 Figure 3M–P
10 Polyalthia verrucipes C. Y. Wu ex P. T. Li XTBG 0020031897(2) Figure 4H
11 Polyalthia laui Merrill SCBG xx240010, xx320026 Figure 4I
12 Polyalthia chinensis S. K. Wu & P. T. Li XTBG 0020023088, 0020150274 Figure 4J
13 Polyalthia yingjiangensis Y. H. Tan and B. Xue XTBG 0020021384(3) Figure 4K
14 Polyalthia obliqua Hook.f. & Thomson XTBG 0020013801(3) Figure 4L
15 Polyalthia longifolia (Sonn.) Thwaites SCBG xx271140, 20055086 Figure 4M
16 Hubera cerasoides (Roxb.) Benth.et Hook.f.ex Bedd. SCBG 20031137(3) Figure 4N
17 Annona squamosa Linn. XTBG 0020071074(3) Figure 5A
18 Annona muricata Linnaeus SCBG 19940242, 20070500 Figure 5B
19 Annona montana Macf XTBG 0019600558A, 2019940014 Figure 5C
20 Annona reticulata L. XTBG 0320140001, 275081 Figure 5D
21 Annona glabra Linn. SCBG xx080063, xx080276, xx080383 Figure 5E
22 Annona cherimola Mill. * XTBG 1520060007(3) Figure 2Q
23 Cananga odorata (Lamarck) J. D. Hooker & Thomson SCBG 20090663, 20140876, xx120033 Figure 4O
24 Cananga odorata var. fruticosa (Craib) J.Sinclair SCBG 20040790(3) Figure 4P
25 Mitrephora thorelii Pierre SCBG 20030719(3) Figure 5F
26 Mitrephora wangii Hu * XTBG 0020022041, 256509, 0019780252 Figure 3A–D
27 Mitrephora teysmannii Scheff SCBG 20042613(3) Figure 5G,H
28 Mitrephora sirikitiae Weeras XTBG 3820130137(3) Figure 5I
29 Pseuduvaria trimera (Craib) YCF Su & RMK. Saunders XTBG 0019880077, 274490, 3020020011 Figure 5J,K
30 Alphonsea monogyna Merrill & Chun SCBG 20011014(3) Figure 4Q
31 Alphonsea mollis Dunn XTBG 0020040356, 0020150149, 286072 Figure 4R
32 Alphonsea glandulosa Y.H. Tan & B. Xue XTBG 0019750173(3) Figure 4S
33 Alphonsea ventricosa (Roxb.) Hook.f.&Thomson XTBG 0019970165(3) Figure 4T
34 Artabotrys hexapetalus (L. f.) Bhanda * XTBG 0020040193, 0020090146 Figure 1I–L
35 Artabotrys hainanensis R. E. Fries SCBG 20012148(3) Figure 5L
36 Artabotrys pilosis Merrill & Chun SCBG 00012542(3) Figure 5M
37 Artabotrys hongkongensis Hance * SCBG 20011052(3) Figure 1M–P
38 Artabotrys pachypetalus B.Xue & Junhao Chen * SCBG 00028772(3) Figure 2A–D
39 Trivalvaria costata (J. D. Hooker & Thomson) I. M. Turner SCBG xx110217(3) Figure 5N,O
40 Trivalvaria carnosa (Teijsm. & Binn.) Scheff XTBG 1320010126(3) Figure 5P
41 Uvaria macrophylla Roxb XTBG 0020023255, 287484, 0020090148 Figure 6A
42 Uvaria grandiflora Roxb * XTBG 3820021106(3) Figure 1A–D
43 Uvaria calamistrata Hance * XTBG 0020201075(3) Figure 2R,S
44 Uvaria tokinensis Finet et Gagnep XTBG 3820020732(3) Figure 6B
45 Uvaria tonkinensis var. subglabra Melodorum SCBG 20030552(3) Figure 6C
46 Uvaria kweichowensis P. T. Li SCBG 20031112(3) Figure 6D
47 Uvaria kurzii (King) P. T. Li * SCBG 042778(3) Figure 1E–H
48 Uvaria boniana Finet et Gagnep SCBG 00044133(3) Figure 6H
49 Uvaria grandiflora var. flava (Teijsm. & Binn.) Scheff XTBG 3820130135(3) Figure 6E
50 Uvaria rufa Bl XTBG 284407(3) Figure 6F
51 Uvaria yunnanensis Hu XTBG 0020070685(3) Figure 5Q
52 Marsypopetalum littorale (Bl.) B. Xue & R. M. K. XTBG 0020012213(3) Figure 5R
53 Goniothalamus chinensis Merr. et Chun * XTBG 3020021381, 0020162454 Figure 3Q–T
54 Goniothalamus calvicarpus Craib * SCBG 20042665, 284928, 275874 Figure 2E–H
55 Goniothalamus gardneri Hook. f. et Thoms * SCBG 20113045(3) Figure 2I–L
56 Goniothalamus saccopetaloides Y.H. Tan and Bin Yang XTBG 3020020407(2) Figure 6G
57 Goniothalamus cheliensis Hu XTBG 3020050005, C30121, 274125 Figure 6K
58 Goniothalamus leiocarpus (W. T. Wang) P. T. Li XTBG 0020013790(2) Figure 6L
59 Goniothalamus howii Merrill & Chun XTBG 0020021240(2) Figure 6I
60 Goniothalamus donnaiensis Finet et Gagnep LMNR AU072086 b Figure 6J
61 Fissistigma wallichii (Hook. f. et Thoms.) Merr XTBG 0020070161, 0020100719 Figure 6P
62 Fissistigma glaucescens (Hance) Merrill SCBG XX271312(2) Figure 6O
63 Fissistigma polyanthum Hook. f. et Thoms * SCBG 20050538(2) Figure 2M–P
64 Fissistigma polyanthoides (Aug. DC.) Merr. XTBG 0020011877, 275869, 275870 Figure 6N
65 Fissistigma acuminatissimum Merrill XTBG 285650(2) Figure 6M
66 Fissistigma bracteolatum Chatt XTBG 374189, 274189, 277352 Figure 6Q
67 Fissistigma maclurei Merr XTBG 0020080638, 286008, 286009 Figure 6R
68 Fissistigma uonicum (Dunn) Merr NMNR 0078738 b Figure 6S
69 Fissistigma thorelii (Pierre ex Finet&Gagnep.) Merr XTBG 0020020480, 0020031109 Figure 6T
70 Meiogyne oligocarpa B. Xue & Y. H. Tan * XTBG 0020013864(2) Figure 3E–H
71 Chieniodendron hainanense (Merr.) Tsiang et P. T. Li SCBG 19980193(2) Figure 5S
72 Cleistopholis glauca Pierre ex Engl. & Diels XTBG 3119800151(2) Figure 5T
73 Orophea hainanensis Merr SCBG 20011196(3) Figure 7A,B
74 Orophea laui Leonardía & Kessler * XTBG 275549, 287159 Figure 3K,L
75 Orophea hirsuta King SCBG 20011196(2) Figure 7C
76 Anaxagorea luzonensis A. Gray DMNR 1020170057(2) Figure 7D
77 Anaxagorea javanica Blume XTBG 3820021060, 0020200920 Figure 7E
78 Miliusa chunii W. T. Wan XTBG 0019970179(2) Figure 7F
79 Miliusa horsfieldii (Bennett) Pierre SCBG 20051897(2) Figure 7G
80 Miliusa sinensis Finet et Gagnep SCBG 011047(2) Figure 7H,I
81 Miliusa chantaburiana Damthongdee & Chaowasku XTBG 0020210589(2) Figure 7J
82 Miliusa glochidioides Hand.-Mazz. SCBG 20113642(2) Figure 7K
83 Miliusa bannaensis X.L. Hou XTBG 0020060634(2) Figure 7L
84 Melodorum fruticosum Lour XTBG 3820021019(2) Figure 7M
85 Melodorum siamense (Scheff.) Bân XTBG 3820021101(2) Figure 7N
86 Disepalum plagioneurum (Diels) D. M. Johnson DMNR 01187407 b Figure 7P
87 Popowia pisocarpa (Bl.) Endl. in Walp. Rep DMNR 0079742 b Figure 7R
88 Asimina triloba Dunal. WBG 20177223(2) Figure 7O,Q
89 Rollinia mucosa (Jacquin) Baillon SCBG AU080617 b Figure 7S
Open in a new tab

Taxoxon with “*” have both types of pollen; others are binucleate. a SCBG (South China Botanical Garden, Chinese Academy of Sciences), XTBG (Xishuangbanna Tropical Botanical Garden of Chinese Academy of Sciences), WBG (Wuhan Botanical Garden Chinese Academy of Sciences), DMNR (Diaoluo Mountain Nature Reserve, Hainan, China), NMNR (Nankun Mountain Nature Reserve, Huizhou, China), LMNR (Longgang Nature Reserve, Guangxi, China). b The voucher column lists the introduction number or specimen number of material used in this study. The numbers in parentheses indicate the number of plants.

Figure 4.

Figure 4

Open in a new tab

Bicellular pollen of Annonaceae shortly before anther dehiscence. (I). (A) Desmos chinensis Lour; (B) Desmos dumosus (roxb.) saff; (C) Desmos yunnanensis (Hu) P. T. Li; (D) Dasymaschalon trichophorum Merr; (E) Desymaschalon filipes (Ridl.) Ridl.Ban; (F) Dasymaschalon macrocalyx Finet & Gagnep; (G) Polyalthia suberosa (Roxburgh) Thwaites; (H) Polyalthia verrucipes C. Y. Wu ex P. T. Li; (I) Polyalthia laui Merrill; (J) Polyalthia chinensis S. K. Wu & P. T. L; (K) Polyalthia yingjiangensis Y. H. Tan & B. Xue; (L) Polyalthia obliqua Hook.f. & Thomson; (M) Polyalthia longifolia (Sonn.) Thwaites; (N) Hubera cerasoides (Roxb.) Benth.et Hook.f.ex Bedd; (O) Cananga odorata (Lamarck) J. D. Hooker & Thomson; (P) Cananga odorata var. fruticosa (Craib) J.Sinclair; (Q) Alphonsea monogyna Merrill & Chun, (R) Alphonsea mollis Dunn; (S) Alphonsea glandulosa Y.H. Tan & B. Xue; (T) Alphonsea ventricosa (Roxb.) Hook.f.&Thomson. Scale bar = 50 μm.

Figure 5.

Figure 5

Open in a new tab

Bicellular pollen of Annonaceae shortly before anther dehiscence. (II). (A) Annona squamosa Linn; (B) Annona muricata Linnaeus; (C) Annona montana Macf; (D) Annona reticulata L; (E) Annona glabra Linn; (F) Mitrephora thorelii Pierre; (G,H) Mitrephora teysmannii Scheff; (I) Mitrephora sirikitiae Weeras; (J,K) Pseuduvaria trimera (Craib) Y. C. F. Su & R. M. K. Saunders; (L) Artabotrys hainanensis R. E. Fries; (M) Artabotrys pilosis Merrill & Chun; (N,O) Trivalvaria costata (J. D. Hooker & Thomson) I. M. Turner; (P) Trivalvaria carnosa (Teijsm. & Binn.) Scheff; (Q) Uvaria yunnanensis Hu; (R) Marsypopetalum littorale (Bl.) B. Xue & R. M. K; (S) Chieniodendron hainanense (Merr.) Tsiang et P. T. Li; (T) Cleistopholis glauca Pierre ex Engl. & Diels. Scale bar = 50 μm.

Figure 6.

Figure 6

Open in a new tab

Bicellular pollen of Annonaceae shortly before anther dehiscence. (III). (A) Uvaria macrophylla Roxb; (B) Uvaria tokinensis Finet et Gagnep; (C) Uvaria tonkinensis var. subglabra Melodorum; (D) Uvaria kweichowensis P. T. Li; (E) Uvaria grandiflora var.flava (Teijsm. & Binn.) Scheff; (F) Uvaria rufa Bl; (G) Goniothalamus saccopetaloides Y.H. Tan & Bin Yang; (H) Uvaria boniana Finet et Gagnep; (I) Goniothalamus howii Merrill & Chun; (J) Goniothalamus donnaiensis Finet et Gagnep; (K) Goniothalamus cheliensis Hu; (L) Goniothalamus leiocarpus (W. T. Wang) P. T. Li; (M) Fissistigma acuminatissimum Merrill; (N) Fissistigma polyanthoides (Aug. DC.) Merr; (O) Fissistigma glaucescens (Hance) Merrill; (P) Fissistigma wallichii (Hook. f. et Thoms.) Merr; (Q) Fissistigma bracteolatum Chatt; (R) Fissistigma maclurei Merr; (S) Fissistigma uonicum (Dunn) Merr; (T) Fissistigma thorelii (Pierre ex Finet&Gagnep.) Merr. Scale bar = 50 μm.

Figure 7.

Figure 7

Open in a new tab

Bicellular pollen of Annonaceae shortly before anther dehiscence. (IV). (A,B) Orophea hainanensis Merr; (C) Orophea hirsuta King; (D) Anaxagorea luzonensis A. Gray; (E) Anaxagorea javanica Blume; (F) Miliusa chunii W. T. Wan; (G) Miliusa horsfieldii (Bennett) Pierre; (H,I) Miliusa sinensis Finet et Gagnep; (J) Miliusa chantaburiana Damthongdee & Chaowasku; (K) Miliusa glochidioides Hand.-Mazz; (L) Miliusa bannaensis X.L. Hou; (M) Melodorum fruticosum Lour; (N) Melodorum siamense (Scheff.) Bân; (O,Q) Asimina triloba Dunal; (P) Disepalum plagioneurum (Diels) D. M. Johnson; (R) Popowia pisocarpa (Bl.) Endl. in Walp. Rep; (S) Rollinia mucosa (Jacquin) Baillon. Scale bar = 50 μm.

Table 2.

Taxon known to have both bicellular and tricellular pollen.

No. Family Taxon % of Anthers with Both Types of Pollen References
1 Annonaceae Uvaria grandiflora Roxb 23.2% (±3%) Present paper
2 Annonaceae Uvaria kurzii (King) P. T. Li 55.0% (±5%) Present paper
3 Annonaceae Uvaria calamistrata Hance 33.3% (±5%) Present paper
4 Annonaceae Annona cherimola Mill 53.0% (±3%) Present paper; [6]
5 Annonaceae Mitrephora wangii Hu 33.3% (±5%) Present paper
6 Annonaceae Mitrephora maingayi Hook. f. et Thoms 33.3% (±5%) [9]
7 Annonaceae Artabotrys hexapetalus (L. f.) Bhandar 25.0% (±5%) Present paper
8 Annonaceae Artabotrys hongkongensis Hance 55.6% (±5%) Present paper
9 Annonaceae Artabotrys pachypetalus B.Xue & Junhao 50.0% (±3%) Present paper
10 Annonaceae Goniothalamus calvicarpus Craib 54.5% (±5%) Present paper
11 Annonaceae Goniothalamus gardneri Hook. f. et Thoms 52.5% (±3%) Present paper
12 Annonaceae Goniothalamus chinensis Merr. et Chun 53.5% (±5%) Present paper
13 Annonaceae Fissistigma polyanthum Hook. f. et Thoms 53.0% (±3%) Present paper
14 Annonaceae Dasymaschalon rostratum Merr. & Chun 50.0% (±2%) Present paper
15 Annonaceae Meiogyne oligocarpa B. Xue & Y. H. Tan 51.0% (±2%) Present paper
16 Annonaceae Orophea laui Leonardía & Kessler 53.0% (±5%) Present paper
17 Annonaceae Polyalthia cheliensis Hu 52.5% (±5%) Present paper
18 Araceae Calla palustris —— [5]
19 Araceae Rhodospatha forgetii —— [5]
20 Araceae Anubias afzelii —— [5]
21 Araceae Dieffenbachia maculata —— [5]
22 Araceae Xanthosoma pilosum —— [5]
23 Araceae Chlorospatha castula —— [5]
24 Araceae Alocasia cuprea —— [5]
25 Asphodelaceae Hemerocallis sp. —— [28]
26 Asteraceae Conyza canadensis (L.) C ronq —— [29]
27 Berberidaceae Leontice incerta Pall. —— [13]
28 Berberidaceae Diphylleia sinensis H. L. Li —— [30]
29 Euphorbiaceae Beyeria leschenaultii —— [4]
30 Gentianaceae Swertia bimaculata —— [31]
31 Gentianaceae Tripterospermum chinense (Migo) Harry Sm. —— [32]
32 Lauraceae Laurelia novae-zelandiae A. Cunn —— [33]
33 Lauraceae Beilschmiedia tara —— [34]
34 Lauraceae Beilschmiedia taw —— [34]
35 Magnoliaceae Michelia figo (Lour.) Spreng. —— [15]
36 Plumbaginaceae Limonium sp. —— [35]
37 Poaceae Bambusa textilis —— [14]
38 Poaceae Shibataea chinensis —— [36]
39 Poaceae Arundinaria simonii f. albostriatus —— [36]
40 Poaceae Pseudosasa viridula —— [12]
41 Poaceae Menstruocalamus sichuanensis —— [36]
42 Poaceae Bambusa multiplex —— [8]
43 Poaceae Sasaella kogasensis ‘Aureostriatus’ —— [11]
44 Poaceae Phyllostachys edulis (Carrière) J. Houzeau —— [16]
45 Ranunculaceae Adonis amurensis Regel et Radde. —— [10]
46 Ranunculaceae Coptis deltoidea C. Y. Cheng et Hsiao —— [37]
47 Saxifragaceae Saxifraga pseudohirculus —— [7]
48 Saxifragaceae Saxifraga caveana —— [7]
49 Solanaceae Solanum phureja —— [38]
50 Solanaceae Solanum japonense Nakai —— [39]
51 Solanaceae Solanum septemlobum Bunge —— [39]
52 Violaceae Viola tricolor L. —— [40]
Open in a new tab

According to the molecular phylogenetic tree of the Annonaceae family constructed by Guo et al. [21], we found that species with both types of pollen are mostly distributed in the relatively evolved tribes, including Annoneae, Uvariaeae (Annonoideae), and Miliuseae (Malmeoideae) (Figure 8). No samples containing both types of pollen were found in the more primitive subfamilies, such as Anaxagoreoideae and Ambavioideae.

Figure 8.

Figure 8

Open in a new tab

The phylogenetic relationships of sampled species at the genus level. Species with both types of pollen are black and marked with *; others are species with binucleate pollen. The phylogenetic tree referenced Guo et al. [21].

4. Discussion

4.1. The Coexistence of Bicellular and Tricellular Pollen May Be More Prevalent in Angiosperms than We Thought

Grayum [5] and Lora et al. [6] reported that the coexistence of bicellular and tricellular pollen occurs transiently prior to dispersal, often with an imbalanced ratio of the two pollen types. If samples are collected prematurely, this mixed population may be misinterpreted as exclusively bicellular pollen. Similarly, incomplete sampling could lead to misclassification as either purely bicellular or tricellular pollen. Such errors have historically caused taxonomic inconsistencies. For example, Calla palustris was described as tricellular by Dudley [41] but bicellular by Brewbaker [3]; subsequent detailed analysis by Grayum [5] revealed that ~5% of pollen completed the second mitotic division prior to dispersal, confirming mixed development. Similarly, while Annonaceae was initially classified as bicellular [3], Rosell et al. [42] identified tricellular pollen in Annona cherimola. later corroborated by Lora et al. [6], who observed both pollen types 9–10 h before anther dehiscence. To mitigate sampling bias, our study exclusively used fully matured flowers at the point of pollen dispersal. We confirmed mixed bicellular and tricellular pollen in Mitrephora maingayi Hook. f. et Thoms [9] and 16 additional species across 10 genera within Annonaceae. This suggests that strict developmental staging of floral material is critical, and we hypothesize that broader sampling with rigorous temporal controls may reveal this phenomenon to be more widespread in angiosperms than currently acknowledged.

4.2. Is the Coexistence of Bicellular and Tricellular Pollen a Special Case or the Third Type?

Previous systematic analysis on pollen cell numbers has treated mixed populations as anomalies of either bicellular or tricellular pollen, often excluding them from samples [1,4,43]. However, within Annonaceae, 17 of 90 species (~19%) and 10 of 30 genera (~33%) exhibit this trait [1,3,22,23,24,25]. Notably, the ratio of bicellular and tricellular pollen is not always imbalanced. A high percentage of both pollen types have been observed in six species of Araceae [5]. Lora et al. [6] reported a mixed population of bicellular (49%) and tricellular (51%) pollen in Annona cherimola Mill. In our study, some anthers contained up to 40% tricellular pollen. In these cases, distinguishing whether this represents a variant of bicellular or tricellular pollen becomes challenging. Thus, at least within Annonaceae, it may be reasonable to take the coexistence of bicellular and tricellular as the third type except for bicellular and tricellular pollen.

In recent years, the discovery of anthers containing both types of pollen has continued to increase in other families, such as the Arundinaria simonii f.albostriatus and Shibataea chinensis Nakai [36], the Bambusa textilis [14], Sasaella kogasensis ‘Aureostriatus’ [11], Bambusa multiplex [8], Pseudosasa viridula [12], Menstruocalamus sichuanensis [36] and Phyllostachys edulis (Carrière) J. Houzeau [16] from Poaceae, the Swertia bimaculata [31] and Tripterospermum chinense (Migo) Harry Sm. [32] from Gentianaceae, the Limonium from Plumbaginaceae [35], which were previously reported as tricellular [3,44]. Similarly, Conyza canadensis (L.) C ronq [29] from Asteraceae, Michelia figo (Lour.) Spreng from Magnoliaceae [15], Coptis deltoidea C. Y. Cheng et Hsiao [37] and Adonis amurensis Regel et Radde [10] from Ranunculaceae, Viola tricolor L. from Violaceae [10], Diphylleia sinensis H. L. Li [30] and Leontice incerta Pall. [12] from Berberidaceae, Solanum japonense Nakai and Solanum septemlobum Bunge from Solanaceae [39], were previously reported as bicellular [1,3]. Additionally, there is also Hemerocallis from Asphodelaceae [28], which had not been reported before. Table 2 lists all the reports of plants shedding both types of pollen. These findings support the idea of treating the coexistence of both types of pollen as the third type of pollen cell number.

4.3. Which Is Primitive and How It Evolved?

The systematic study of pollen cell numbers has been developed and controversial for over a century. Schürhoff [45] firstly observed that most plant families produce either bicellular or tricellular pollen. Schnarf [46,47,48] and Brewbaker [3] further noted that taxa with bicellular pollen were predominantly basal within the phylogenetic tree. Webster and Rupert [4] proposed the “Schiirhoff-Brewbaker Law”, which claims that bicellular pollen was primitive in the angiosperms as a whole and that the evolution process from bicellular to tricellular pollen was irreversible. However, this hypothesis was challenged by Gardner [34], who identified tricellular pollen in Lauraceae (a primitive group), and Webster et al. [49], who found tricellular pollen in early-diverging Euphorbieae. Williams et al. [1] analyzed 2511 species and modeled trait evolution using time-calibrated phylogenines, revealing reversible transitions between bicellular and tricellular states, as well as differential diversification rates between the two lineages. Although they questioned the likelihood of a tricellular origin, they could not conclusively determine the ancestral state.

Interestingly, gymnosperms exhibit varying mitotic divisions during pollen maturation, releasing pollen at different male gametophyte developmental stages [50]. However, previous systematic studies on angiosperms pollen have only considered strictly bicellular and tricellular forms, often disregarding mixed cases as anomalies. If the coexistence of both types of pollen are taken into account, where should it be located? From the limited known results (Table 2), the coexistence of both types of pollen are found in primitive families such as Lauraceae, Magnoliaceae, Annonaceae, and relatively advanced families such as Asteraceae, Gentianaceae and Violaceae. However, in Annonaceae, the coexistence of both types of pollen are mostly distributed in relatively advanced tribes such as Annoneae, Uvariaeae (Annonoideae), and Miliuseae (Malmeoideae) (Figure 8). Lora et al. [6] indicated that the coexistence of both types of pollen may have stronger adaptability than bicellular and tricellular pollen because the environmental conditions of pollen after dispersal are unpredictable, and the combined type of pollen could increase the chance of fertilization. Thus, the coexistence of both types of pollen may be advanced.

Franchi et al. [51] proposed that pollen, like seeds, may undergo developmental arrest (DA) under unfavorable conditions. DA in pollen refers to the phenomenon where the development of pollen slows or temporarily halts before reaching full maturity. This process typically occurs at the bicellular stage, during which the pollen consists of one vegetative cell and one generative cell that has not yet divided into two sperm cells, such as drought conditions Furthermore, DA can occur at any stage of pollen development; it might manifest as bicellular pollen when arrested at the bicellular stage, as either type of pollen when halted during the transition from bicellular to tricellular stage, or as tricellular pollen when environment conditions are favorable and no DA takes place. DA in pollen is strongly associated with the acquisition of desiccation tolerance (DT), which extends pollen viability during air travel [52]. Williams and Brown [53] found a close relationship between the number of pollen cells and pollen water content, revealing that most tricellular pollen exhibited a relatively high water content, whereas bicellular pollen showed comparatively low water content. From the perspective of DA, bicellular pollen appears to represent an evolutionary adaptation from aquatic to terrestrial plants. Additionally, the retrogradation of tricellular pollen observed by Williams et al. [1] may have resulted from a loss of dehydration tolerance during subsequent evolutionary processes.

5. Conclusions

In the present study, we identified 16 species from 10 genera of Annonaceae that shed both types of pollen. Including reports from other families, approximately 15 families, 40 genera, and 52 species are known to produce both types of pollen. The coexistence of bicellular and tricellular pollen might be the third type of pollen cell number. And the systematic relationship among them is needed to be reanalyzed.

Acknowledgments

The authors thank Peng Caixia, Chen Ling and Xue Bine, Liao Jingping, Wen bin, Lai Han and Deng Xingmin for their assistance with material collection.

Author Contributions

Conceptualization, Y.G.; methodology, Y.G., J.P. and Q.Z.; software, Q.Z.; validation, C.X.; formal analysis, Y.G. and J.P.; investigation, Y.G. and J.P.; resources, Q.Z. and C.X.; data curation, Y.G.; writing—original draft preparation, Y.G.; writing—review and editing, J.P.; visualization, Q.Z.; supervision, Y.G.; project administration, Y.G.; funding acquisition, Y.G. All authors have read and agreed to the published version of the manuscript.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The original contributions presented in this study are included in the article. Further inquiries can be directed at the corresponding author.

Conflicts of Interest

The authors declare no conflicts of interest.

Funding Statement

This research was funded by the National Natural Sciences Foundation of China (grant number 31800184) and the “Jinying Star” Talent Project of Guangdong Academy of Agricultural Sciences (grant number R2023PY-JX021).

Footnotes

Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

References

  • 1.Williams J.H., Taylor M.L., O’Meara B.C. Repeated evolution of tricellular (and bicellular) pollen. Am. J. Bot. 2014;101:559–571. doi: 10.3732/ajb.1300423. [DOI] [PubMed] [Google Scholar]
  • 2.Schurhoff P.N. Die Zytologie der Blütenpfl Anzen. Enke; Stuttgart, Germany: 1926. [Google Scholar]
  • 3.Brewbaker J.L. Distribution and phylogenetic significance of bicellular and tricellular pollen grains in angiosperms. Am. J. Bot. 1967;54:1069–1083. doi: 10.1002/j.1537-2197.1967.tb10735.x. [DOI] [Google Scholar]
  • 4.Webster G.L., Rupert E.A. Phylogenetic significance of pollen nuclear number in Euphorbiaceae. Evolution. 1973;27:524–531. doi: 10.2307/2407315. [DOI] [PubMed] [Google Scholar]
  • 5.Grayum M.H. Phylogenetic implications of pollen nuclear number in the Araceae. Plant Syst. Evol. 1986;151:145–161. doi: 10.1007/BF02430271. [DOI] [Google Scholar]
  • 6.Lora J., Herrero M., Hormaza J.I. The coexistence of bicellular and tricellular pollen in Annona cherimola (Annonaceae): Implications for pollen evolution. Am. J. Bot. 2009;96:802–808. doi: 10.3732/ajb.0800167. [DOI] [PubMed] [Google Scholar]
  • 7.Zhang Z.X., Gornall R.J. Systematic significance of pollen nucleus number in the genus Saxifraga (Saxifragaceae) Plant Syst. Evol. 2011;295:13–22. doi: 10.1007/s00606-011-0456-9. [DOI] [Google Scholar]
  • 8.Lin S., Li J., Zhao R., Dong X., Ding Y. The development of flowering bud differentiation and male gametophyte of Bambusa multiplex. J. Nanjing For. Univ. (Nat. Sci. Ed.) 2015;39:51–56. (In Chinese) [Google Scholar]
  • 9.Gan Y.Y., Xu F.X. The coexistence of bicellular and tricellular pollen. Grana. 2018;58:129–132. doi: 10.1080/00173134.2018.1509234. [DOI] [Google Scholar]
  • 10.Sun Y., Wang L., Yang X., He M. Megasporogenesis, microsporogenesis and development of male anda female gametophytes of Adonis amurensis Regel et Radde. J. Northest For. Univ. 2017;45:61–65. [Google Scholar]
  • 11.Wang X., Jiang M.Y., Lin S.Y., Yao W.J., Ding Y.L. Inflorescence architecture of male and female gametophytes of Sasaella kogasensis ‘Aureostriatus’. J. Anhui Agric. Univ. 2021;48:9–14. (In Chinese) [Google Scholar]
  • 12.Zhao W.Q., Guo C.C., Wang Y., Xiao J., Wu Z.C., Yang G.Y., Yu F., Liu H.W. Sporogenesis and gametogenesis of Pseudosasa viridula. Guihaia. 2022;42:143–151. (In Chinese) [Google Scholar]
  • 13.Zhang J., Ma H. Cyto-embryological analysis of wild kentucky bluegrass germplasm in Gansu Province, China. Agronomy. 2023;13:1569. doi: 10.3390/agronomy13061569. [DOI] [Google Scholar]
  • 14.Yan Y.D., Zhou L., Wang S.G. Floral morphology and development of female and male gametophytes of Bambusa textilis. For. Res. 2024;37:150–158. (In Chinese) [Google Scholar]
  • 15.Liu S., Nie T., Wu Q., Jin L., Wan X., Yin Z. The basic characteristics of stamen morphogenesis and development in Michelia figo. J. Nanjing For. Univ. (Nat. Sci. Ed.) 2024;48:34–40. (In Chinese) [Google Scholar]
  • 16.Zhou J.M., Sun L.F., Feng Y., Lian C., Ran H., Zhang Y., Guo Q. Observation on anther development of Phyllostachys edulis. Guihaia. 2016;36:231–235. (In Chinese) [Google Scholar]
  • 17.Chawasku T., Kessler P.J.A., van der Ham R.W.J.M. A taxonomic revision and pollen morphology of the genus Dendrokingstonia (Annonaceae) Bot. J. Linn. Soc. 2012;168:76–90. doi: 10.1111/j.1095-8339.2011.01187.x. [DOI] [Google Scholar]
  • 18.Azeez S.O., Folorunso A.E. Phenology and pollen studies of some species of Annonaceae in Nigeria. IFE J. Sci. 2014;16:171–178. [Google Scholar]
  • 19.Gan Y.Y., Liu Y., Xu F.X. Pollen morphology of selected species from Annonaceae. Grana. 2015;54:271–281. doi: 10.1080/00173134.2015.1096302. [DOI] [Google Scholar]
  • 20.Shao Y.Y., Xu F.X. Studies on pollen morphology of selected species of Annonaceae from Thailand. Grana. 2017;1:161–177. doi: 10.1080/00173134.2017.1350204. [DOI] [Google Scholar]
  • 21.Guo X., Tang C.C., Thomas D.C., Couvreur T.L., Saunders R.M. A mega-phylogeny of the Annonaceae: Taxonomic placement of five enigmatic genera and support for a new tribe, Phoenicantheae. Sci. Rep. 2017;7:7327. doi: 10.1038/s41598-017-07252-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.Periasamy K., Swamy B.G.L. Studies in the Annonaceae I. Microsporogenesis in Cananga odorata and Miliusa wightiana. Phytomorphology. 1959;9:251–263. [Google Scholar]
  • 23.Periasamy K., Kandasamy M.K. Development of the anther of Annona squamosa L. Ann. Bot. 1981;48:885–893. doi: 10.1093/oxfordjournals.aob.a086196. [DOI] [Google Scholar]
  • 24.Periasamy K., Thangavel S. Anther development in Xylopia nigricans. Proc. Plant Sci. 1988;98:251–255. doi: 10.1007/BF03053795. [DOI] [Google Scholar]
  • 25.Tsou C.H., Fu Y.L. Octad pollen formation in Cymbopetalum (Annonaceae): The binding mechanism. Plant Syst. Evol. 2007;263:13–23. doi: 10.1007/s00606-006-0471-4. [DOI] [Google Scholar]
  • 26.Lora J., Herrero M., Hormaza J.I. Pollen performance, cell number, and physiological state in the early-divergent angiosperm Annona cherimola Mill (Annonaceae) are related to environmental conditions during the final stages of pollen development. Sex. Plant Reprod. 2012;25:157–167. doi: 10.1007/s00497-012-0187-2. [DOI] [PubMed] [Google Scholar]
  • 27.Fu L., Zeng Q.W., Liao J.P., Xu F.X. Anatomy and ontogeny of unisexual flowers in dioecious Woonyoungia septentrionalis (Dandy) Law (Magnoliaceae) J. Syst. Evol. 2009;47:263–272. doi: 10.1111/j.1759-6831.2009.00029.x. [DOI] [Google Scholar]
  • 28.Cheng X.J., Wang R.M., Zhai B.J., Gong F.F., Hou F.F., Wang J.Y., Wu J., Xing G.M., Kang X.P., Li S. Preliminary study on correlation between microspore development and bud length of Hemerocallis and anther culture. J. Shanxi Agric. Sci. 2021;49:170–174. (In Chinese) [Google Scholar]
  • 29.Huang H.Y., Long H., Li P. Studies on embryology of Conyza canadensis (L.) C ronq. J. Wuhan Bot. Res. 2009;37:233–241. (In Chinese) [Google Scholar]
  • 30.Huang H.Y., Long H., Li P. Genesis of microspore, megaspore and the development of male gametophyte, female gametophyte in Diphylleia sinensis. Guihaia. 2010;30:36–44. (In Chinese) [Google Scholar]
  • 31.Huang H.Y., Long H., Ting-Ting Y.I., Li L.I. Microsporogenesis and the development of male gametopyte in Swertia bimaculata. Guihaia. 2010;30:584–593. (In Chinese) [Google Scholar]
  • 32.Huang H.Y., Long H., Yi T.T., Li P. Genesis of microspore and the development of male gametophyte in Tripterospermum discoideum. Guihaia. 2010;30:316–323. (In Chinese) [Google Scholar]
  • 33.Sampsom F.B. Studies on the Monimiaceae III. Gametophyte development of Laurelia novae-zelandiae A. Cunn. (subfamily Antherospermoideae) Aust. J. Bot. 1969;17:425–439. doi: 10.1071/BT9690425. [DOI] [Google Scholar]
  • 34.Gardner R.O. A survey of the distribution of bicellular and tricellular pollen in the New Zealand flora. N. Z. J. Bot. 1975;13:361–366. doi: 10.1080/0028825X.1975.10430333. [DOI] [Google Scholar]
  • 35.Róis A.S., Teixeira G., Sharbel T.F., Fuchs J., Martins S., Espírito-Santo D., Caperta A.D. Male fertility versus sterility, cytotype, and DNA quantitative variation in seed production in diploid and tetraploid sea lavenders (Limonium sp., Plumbaginaceae) reveal diversity in reproduction modes. Sex. Plant Reprod. 2012;25:305–318. doi: 10.1007/s00497-012-0199-y. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 36.Lin S.Y. Ph.D. Thesis. Nanjing Forestry University; Nanjing, China: 2009. Studies on the Reproductive Biology of Shibataea chinensis and Arundinaria simoniif Albostriatus. (In Chinese) [Google Scholar]
  • 37.Zhu M. Cheng et Hsiao and Coptis Chinensis Franch. Chengdu University of Tranditional Chinese Medicine; Chengdu, China: 2011. Studies on Comparative Embryology Between Coptis deltoidea C. Y. [Google Scholar]
  • 38.Dnyansagar V.R., Cooper D.C. Development of the seed of Solanum phureja. Am. J. Bot. 1960;47:176–186. doi: 10.1002/j.1537-2197.1960.tb07111.x. [DOI] [Google Scholar]
  • 39.Liu Y.S., Gu L., Liu J.X. Sporogenesis, gametogenesis and pollen morphology of Solanum japonense and S. septemlobum (Solanaceae) Arch. Biol. Sci. 2018;70:313–320. doi: 10.2298/ABS170724048L. [DOI] [Google Scholar]
  • 40.Wang X.L., Xu Y.Q., Wang Y., Hu B.Z. Anatomical studies on microsporogenesis and development of male gametophyte in pansy. J. Northeast Agric. Univ. 2014;45:60–65. (In Chinese) [Google Scholar]
  • 41.Dudley M.H. Morphological and cytological studies of Calla palustris. Bot. Gaz. 1937;98:556–571. doi: 10.1086/334659. [DOI] [Google Scholar]
  • 42.Rosell P., Herrero M., Sauco V.G. Pollen germination of cherimoya (Annona cherimola Mill.). In vivo characterization and optimization of in vitro germination. Sci. Hortic. 1999;8:251–265. doi: 10.1016/S0304-4238(99)00012-6. [DOI] [Google Scholar]
  • 43.Schürhoff P.N. Die Zytologie der Blütenpflanzen. Enke; Stuttgart, Germany: 1936. [Google Scholar]
  • 44.Ma W.L. Botany. Higher Education Press; Beijing, China: 2022. [Google Scholar]
  • 45.Schurhoff P.N. Zytologische Untersuchungen in der Reihe Geraniales. Jahrb. Wiss. Bot. 1924;63:707–758. [Google Scholar]
  • 46.Schnarf K. Vergleichende Embryologie der Angiospermen. Gebrlider Borntraeger; Berlin, Germany: 1931. [Google Scholar]
  • 47.Schnarf K. Studien iiber den Bau der Pollenkorner der Angiospermen. Planta. 1937;27:450–465. doi: 10.1007/BF01917034. [DOI] [Google Scholar]
  • 48.Schnarf K. Variation im Bau der Pollenkorner der Angiospennen. Tabulae Biol. 1939;17:72–89. [Google Scholar]
  • 49.Webster G.L., Rupert E., Koutnik D. Systematic significance of pollen nuclear number in Euphorbiaceae, tribe Euphorbieae. Am. J. Bot. 1982;69:407–415. doi: 10.1002/j.1537-2197.1982.tb13274.x. [DOI] [Google Scholar]
  • 50.Fernando D.D., Quinn C.R., Brenner E.D., Owens J.N. Male gameto-phyte development and evolution in extant gymnosperms. Int. Plant Dev. Biol. 2010;4:47–63. [Google Scholar]
  • 51.Franchi G.G., Piotto B., Nepi M., Baskin C.C., Baskin J.M., Pacini E. Pollen and seed desiccation tolerance in relation to degree of developmental arrest, dispersal, and survival. J. Exp. Bot. 2011;62:5267–5281. doi: 10.1093/jxb/err154. [DOI] [PubMed] [Google Scholar]
  • 52.Pacini E., Dolferus R. Pollen developmental arrest maintaining pollen fertility in a world with a changing climate. Front. Plant Sci. 2019;10:679. doi: 10.3389/fpls.2019.00679. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 53.Williams J.H., Brown C.D. Pollen has higher water content when dispersed in a tricellular state than in a bicellular state. Acta Bot. Bras. 2018;32:454–461. doi: 10.1590/0102-33062018abb0129. [DOI] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Data Availability Statement

The original contributions presented in this study are included in the article. Further inquiries can be directed at the corresponding author.

Articles from Biology are provided here courtesy of Multidisciplinary Digital Publishing Institute (MDPI)

RESOURCES